Drinking water treatment drastically modifies multi-kingdom water communities, but clinically relevant opportunistic bacterial pathogens persist

Controlling pathogenic microorganisms during water treatment processes is a major public health concern. However, the impact of different water sources and treatment processes on the drinking water microbiome and opportunistic bacterial pathogens remains poorly understood. Here, we report the analyses of two water systems: a river and groundwater source, and their corresponding treated waters, sampled bi-weekly for 6 months. Microbial communities were analysed through a metabarcoding and dPCR workflow coupling universal prokaryotic and eukaryotic primers with genus-targeting primers for Legionella and Pseudomonas to achieve a deep taxonomic resolution. We show that microbial communities in groundwater were more diverse than those in river water and that water treatment affected -diversity. Water treatment had a stronger influence on community composition than water system or seasonal variation. Despite lower bacterial loads, the relative abundance of Legionella, Pseudomonas, and non-tuberculous Mycobacteria (NTM) remained stable or even increased after water treatment, suggesting that conventional treatment processes promote enrichment of specific taxa. Excitingly, the genus-targeting analyses revealed a remarkable and previously uncharacterized diversity of Legionella species. L.pneumophila, pathogenic Pseudomonas clades and P. aeruginosa were detected in all environments, emphasizing their ability to adapt to diverse ecological conditions. Analysis of the eukaryotic fraction emphasized the persistence and emergence of eukaryotic hosts associated with these waterborne opportunistic pathogens. Our findings offer important insights into how water source characteristics and treatment processes influence the composition of the drinking water microbiome, while the high taxonomic resolution achieved for waterborne opportunistic bacterial pathogens underscores their ecological adaptability.